Breast cancer (BC) incidence rates are much higher in post- than pre-menopausal women. Estrogen is a well-known risk factor for BC development. Yet, total estrogen levels are lower in post- than pre-menopausal women. This apparent discrepancy calls for new research to better understand BC etiology and mechanisms. In the present study, we hypothesize that an accumulation of high iron levels due to menopause is one of the pre-neoplastic changes in post-menopausal women, and in the presence of high local estrogen in the breast tissue, both iron and estrogen greatly promote breast tumor growth and oxidative damage. We will test this hypothesis using an ovariectomized (OVX) transgenic iron-loaded mouse model carrying hemochromatosis (HFE) gene mutation. This model has a decreased estrogen level because of the OVX, but an increased iron level due to the HFE gene mutation, thus mimicking post-menopausal conditions. In Aim1, we will assess the role of estrogen in mammary epithelial proliferation and spontaneous tumor formation in wild-type and iron loaded HFE+/- mice. Female wild-type C57BL6 mice and HFE+/- mice will undergo OVX. After implantation of 17beta-estradiol (E2), Ki-67, a proliferation-specific antigen, and spontaneous tumor incidence of the iron-loaded mutant and wild-type mice will be compared. In Aim 2, we will assess whether E2 induces a higher tumor take and growth of mammary cancer in HFE+/- mice than in wild-type mice. MXT+ mammary cancer cells will be inoculated into the mammary fat pads of wild-type and HFE+/- mice. Mammary cancer tumorigenicity and tumor latency as well as tumor volume will be measured. Levels of oxidative DNA and protein in breast tissues will be determined and compared. In Aim 3, we will determine whether E2-mediated tumor growth is iron-dependent. The MXT+ cell line will be transfected with small interfering RNA (siRNA) of transferring receptor. Mammary cancer will be measured and compared between the siRNA transfectants and the parental cells. We expect to show that iron plays an important role in estrogen-dependent BC and this new direction of research may greatly benefit the health of female baby- boomers. [unreadable] [unreadable] [unreadable]